LeukoVantage^® Myeloid Neoplasm Mutation Panels

These LeukoVantage panels detect myeloid neoplasm-associated mutations in 48 genes associated with acute myeloid leukemia (AML), myelodysplastic syndromes (MDS), and/or myeloproliferative neoplasms (MPNs) (Table). They can be used for diagnosis, prognosis, and monitoring of these 3 diseases.

The following factors were considered when choosing genes for this panel:

• Frequency of mutation in myeloid neoplasms
• Ability to indicate a distinct AML, MDS, or MPN subtype
• Strength of genetic association with hematopoietic function, myeloid transformation, or leukemic progression

Results are reported in a tabular format with the following individual tables:

• Overall summary
• Biomarker summary
• Biomarker interaction
• Variants of unknown significance
• Biomarkers with no alterations
• Regions with poor sequence quality
• Biomarker details with clinical annotation

Reporting Workflow: Only mutations present in the interrogated regions of the genes are reported. Normal population variations and single-nucleotide polymorphisms (SNPs), promoter and intronic variations (except splice variants), synonymous variants, and benign variants are not included in the report. This test can detect single-nucleotide variants (SNVs) and insertion and deletion (INDEL) variants present at >5% to 10% variant frequency. Insertion or deletion detection is limited to 33 bp for insertions and 52 bp for deletions (insertions or deletions outside these limits may be reported at the medical director’s discretion). Once clinically relevant alterations are detected, an XML file is generated that includes de-identified patient information. The XML file is transmitted securely to a third-party annotation group for clinical annotation. The information transmitted includes the disease (eg, AML), molecular alterations identified (eg, KRAS G12A; IDH1 R132C), and nucleotide position(s) of the alteration(s). The clinical annotation includes information regarding FDA-approved drugs for the indication, National Comprehensive Cancer Network (NCCN) Guidelines^® information, clinical trials, and, if available, prognostic information relevant to the indication and molecular alterations detected. The information is then parsed and automatically uploaded into case review and sign-out portal. A pathologist/director reviews the results and the clinical annotation before the reports are finalized.

Yes. Leukemia-associated fusion transcripts arising from chromosomal translocations, such as BCR-ABL1 and PML-RARA, are not included in the LeukoVantage panels.

Testing for these translocations is available using (1) break-apart or fusion FISH probes; or (2) highly sensitive, quantitative, reverse-transcription−polymerase chain reaction (PCR) assays. Because of their specificity, broad dynamic range, and high analytic sensitivity, PCR methods are preferred for monitoring most common leukemia translocations. This is particularly true for BCR-ABL1 gene rearrangements, for which transcript levels are linked to therapeutic milestones.

The LeukoVantage AML panel can be used to assess AML subclass and prognosis based on the presence of recurrent genetic abnormalities. These include variants in NPM1, CEBPA, and RUNX1.¹

A number of genes are cited in guidelines (NCCN^® and World Health Organization[WHO]) for their prognostic importance.^1,2 The panel identifies the most important gene-based AML prognostic factors, including mutations in ASXL1, FLT3, KIT, RUNX1, TET2, and TP53.^1-4

In addition, there are a growing number of genes for which there are targeted therapies, including FLT3, IDH1/2, and JAK2. Several others are also in the pharmaceutical pipeline.^4,5

The panel can be used to

• Identify mutations that enable a definitive diagnosis of MDS in the presence of clonal hematopoiesis. This is useful for patients with persistent cytopenias and evidence of morphological dysplasia, but with no cytogenetic abnormalities. This is particularly common in patients with early MDS who may have subtle morphologic disturbances.⁶

• Expand prognostic capability for patients with established MDS by identifying relevant gene mutations.⁷

This panel can assist in the diagnosis and subclassification of MPN. For MPN, primary driver mutations in certain genes satisfy diagnostic criteria for MPN (eg, JAK2 V617F or exon 12 mutation in polycythemia vera). However, in many WHO diagnostic criteria, even mutations in secondary drivers can be used as evidence to support a diagnosis in the context of other supporting information.¹ In this way, any of the genes in the test can have diagnostic significance in MPNs. In addition, many of the secondary driver mutations confer prognostic information.^1,8

This panel can detect mutations in growth-promoting genes, such as KRAS and NRAS, and in epigenetic regulators, such as ASXL1, CBL, EZH2, KDM6A, and TET2. Knowledge of these mutations can help in establishing a definitive diagnosis and subclassification in cases lacking BCR-ABL1 and mutations in JAK2, CALR, CSF3R, and MPL.^8,9

This 47-gene panel provides a more comprehensive review of the mutation profile, which may be preferable when the category of the myeloid neoplasm has not yet been established or it has features that overlap with AML, MDS, or MPNs.

These NGS assays are performed using DNA bait capture methodology on the NextSeq^® (Illumina^®) platform. They detect mutations associated with AML, MDS, and MPNs. Within the regions interrogated, SNVs, and small INDELs are detected.

Following DNA extraction from leukocytes, a targeted, amplicon-based NGS method is used to detect mutations. The lower limit of sensitivity is 5% (including MLL PCR), which is superior to that of traditional Sanger DNA sequencing. The percentage of mutation reads will be reported and can be used to assess the size of the clonal population. However, these tests have not been specifically evaluated for assessing minimal residual disease.

Results can be expected in 14 days after sample receipt.